摘要
目的探讨抑制miR-301a对胶质瘤U87细胞增殖、凋亡和侵袭的影响。方法体外培养人脑胶质瘤U87细胞,将细胞分为空白对照组、阴性转染组(阴性序列转染U87细胞)、miR-301a抑制剂转染组(miR-301a抑制剂转染U87细胞)。采用MTT法检测各组细胞的增殖;平板细胞克隆形成实验检测菌落形成;划痕实验检测细胞迁移能力;Transwell法检测细胞迁移、侵袭能力;流式细胞仪检测细胞的凋亡率;RTPCR法检测细胞中miR-301a、PTEN mRNA表达;Western blot检测细胞PTEN、B淋巴细胞瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)、Caspase-9蛋白表达。结果与空白对照组、阴性转染组比较,miR-301a抑制剂转染组转染后不同时间点的细胞抑制率均升高,呈时间依赖性(均P<0.05)。miR-301a抑制剂转染组的菌落数量明显降低,迁移距离短,细胞迁移、侵袭数量明显降低,细胞凋亡率升高(均P<0.05);miR-301a mRNA表达水平降低,PTEN mRNA表达水平升高;PTEN、Bax、Caspase-3、Caspase-9蛋白表达水平明显升高,Bcl-2蛋白表达水平降低(均P<0.05)。结论抑制miR-301a的表达,可抑制胶质瘤U87细胞的增殖、迁移、侵袭能力,促进胶质瘤U87细胞凋亡。
Objective To explore the effects of inhibition of miR-301 a on the proliferation,apoptosis and invasion of glioma U87 cells.Methods Human glioma U87 cells were cultured in vitro,and the cells were divided into blank control group,negative transfection group（negative sequence transfected U87 cells）,and miR-301 a inhibitor transfection group（miR-301 a inhibitor transfected U87 cells）.The cell proliferation of each group was detected by MTT.The cell clone and formation experiment on the flat was used to detect the colony formation.The migration ability of cells was detected by scratch test and the cell migration and invasion abilities were detected by Transwell.Flow cytometry was used to detect the cell apoptosis.The expression of miR-301 a and PTEN mRNA in the cells was detected by RT-PCR.Western Blot was used to detect the expressions of PTEN,B lymphocyte tumor-2（Bcl-2）,Bcl-2 related X protein（Bax）,Caspase-3 and Caspase-9 in cells.Results Compared with the blank control group and the negative transfection group,the cell inhibition rate of the miR-301 a inhibitor transfected group at different time points increased,and in a time dependent（all P 〈 0.05）.Compared with the blank control group and the negative transfection group,the number of colonies in the miR-301 a inhibitor transfected group decreased and the migration distance decreased（all P 〈 0.05）.Compared with the blank group and negative transfection group,the numbers of the cell migration and invasion in the miR-301 a inhibitor transfected group decreased and the rate of cell apoptosis increased（all P 〈 0.05）.Compared with the blank group and negative transfected group,the expression of miR-301 a mRNA in the miR-301 a inhibitor transfected group decreased and the expression of PTEN mRNA increased（all P 〈0.05）.The expression of PTEN,Bax,Caspase-3 and Caspase-9 increased and the expression of Bcl-2 protein decreased（P 〈 0.05）.Conclusion Inhibition of miR-301 a can inhibit the proliferation,migration and invasion of glioma U87 cells,and promote the apoptosis of glioma U87 cells.
作者
罗安志
程敬民
LUO Zhi-an;CHEN Jing-ming(Department of Neurosurgery,Meishan City People’s Hospital,Meishan 620020,Chin)
出处
《临床神经外科杂志》
CAS
2018年第4期255-260,共6页
Journal of Clinical Neurosurgery
基金
2012四川省卫生厅科研立项课题(120332)
